Armadillo (arm) repeat is a 42 amino acid protein—protein interaction motif (Peifer et al., 1994; Hatzfeld, 1999; Andrade et al., 2001). The repeat was first identified in the Drosophila segment polarity gene armadillo (Riggleman, 1989) and since then in many eukaryotic proteins involved in cell signaling or cellular architecture. Armadillo and its vertebrate homolog β-catenin are components of the Wingless and the Wnt signaling pathways, which determine the pattering of Drosophila embryo body segments and vertebrate cell fates, respectively (Polakis, 2000). When triggered by the Wingless or Wnt growth factor signal, otherwise unstable armadillo/β-catenin becomes stabilized, translocates into the nucleus, and, together with the TCF/LEF subfamily of transcription factors, activates the Wingless/Wnt target genes. β-catenin also plays a structural role in cell—cell adhesion, by linking the transmembrane adhesion molecules cadherins to actin cytoskeleton.
Pfam (http://www.sanger.ac.uk/Software/Pfam/) and SMART (http://smart.embl-heidelberg.de/) protein databases enlist more than 90 Arabidopsis arm repeat proteins. Based on their sequence homology, these proteins can be grouped into several different subfamilies such as impotin-α, kinesin, and U-box protein families (Coates, 2003). However, the functions of the Arabidopsis and other plant arm repeat proteins have not been characterized in detail except those of ARC1 and PHOR1. ARC1 interacts with an S-locus receptor kinase of Brassica (Gu et al., 1998) and has been demonstrated to be a positive regulator of the self-incompatibility response (Stone et al., 1999). A recent study shows that ARC1 promotes ubiquitination and proteasomal degradation of compatibility factors in pistil (Stone et al., 2003). The potato arm repeat protein PHOR1, on the other hand, is involved in gibberellin (GA) signaling (Amador et al., 2001). Antisense suppression of its expression reduces GA sensitivity and plant height, whereas its overexpression increases GA sensitivity and internode length.
BTB (BR-C, ttk, and bab) domain is another evolutionarily conserved protein—protein interaction domain (Bardwell and Treisman, 1994; Zollman et al., 1994). The ˜120 amino acid motif, also known as POZ (poxvirus and zinc finger) domain, was first identified in a group of poxvirus proteins and in Drosophila zinc finger proteins, Broad-Complex (BR-C), Tramtrak (Ttk), and Bric-a-brac (bab). Subsequently, it has been found that the BTB/POZ domain is present in 5–10% of zinc finger transcription factors and in some actin-binding proteins or ion channels (Aravind and Koonin, 1998; Collins et al., 2001). Arabidopsis genome contains approximately 80 BTB domain proteins. However, only three of them have been reported to date: NPH3 and RPT2, signal transducers of phototrophic response (Motchoulski and Liscum, 1999; Sakai et al., 2000), and NPR1/NIM1, a regulator of gene expression during systemic acquired response (Cao et al., 1997; Ryals et al., 1997).
Plant hormone abscisic acid (ABA) controls various aspects of plant growth and development (Finkelstein et al., 2002). It inhibits germination and postgermination growth at high concentrations, although it is necessary for normal seedling growth. It regulates seed maturation process and prevents embryos from precocious germination. During vegetative growth, ABA plays an essential role in adaptation to various abiotic stresses such as drought, high salinity and cold (Xiong et al., 2002). Extensive genetic and biochemical studies have been done to identify the regulatory components of various aspects of ABA response. As a consequence, a large number of ABA signaling components have been reported that include transcription factors, kinases/phosphatases, RNA-binding proteins, G-proteins, and secondary messengers (Finkelstein et al., 2002; Xiong et al., 2002).
During vegetative growth, ABA controls the expression of numerous genes associated with adaptive responses to drought and other abiotic stresses (Ramanulu and Bartels, 2002; Shinozaki et al., 2003). The ABA-regulation of stress-responsive genes is largely mediated by cis-regulatory elements sharing the CACGTGGC consensus. Previously, we and others identified a small subfamily of basic leucine zipper (bZIP) class transcription factors that interact with the elements (Choi et al., 2000; Uno et al., 2000). We subsequently showed that the factors, named as ABFs (i.e., ABF1–ABF4) or AREBs (i.e., AREB1–AREB3), are involved in ABA and various abiotic stress responses (Kang et al., 2002; Kim et al., 2004). In particular, ABF2/AREB1, which will be referred to as ABF2 hereafter, regulates seedling growth rate and plays an essential role in glucose-induced developmental arrest process. Its overexpression phenotypes such as altered ABA sensitivity and multiple stress tolerance also suggest that it is involved in ABA and stress responses. We are interested in delineating the ABA signaling pathway(s) that lead to the ABF-dependent ABA/stress-responsive gene expression in vegetative tissues. Toward this end, we carried out two-hybrid screens to isolate proteins that interact with ABF family members to modulate their activities. Here, we describe an arm repeat and BTB/POZ domain protein that interacts with ABF2. In vivo analysis of its function revealed that the ABF2-interacting protein is a novel ABA signaling component that regulates seed germination, seedling growth, glucose response, and ABA/stress responses. In particular, overexpression of ARIA in Arabidopsis results in enhanced seedling survival under high salt conditions, indicating that it can be utilized to develop salt-tolerant plants.